Alcohol addiction is one of the most common addictive disorders with a high rate of relapse among recovering alcoholics. The development of addiction and subsequent relapse following abstinence have been linked to several of the processes and neural structures that contribute to learning and memory, and multiple lines of research suggest that alcohol addiction may be a form of maladaptive learning. Ethanol withdrawal severity is an indicator of addiction development and a major factor in relapse. Our lab recently identified Kcnj9 as a quantitative trait gene (QTG) for ethanol withdrawal in mice. Kcnj9 codes for the GIRK3/Kir3.3 subunit of the G- protein coupled inwardly rectifying potassium (GIRK) channel family, and GIRK3 knockout (KO) and heterozygote (HET) mice show less severe ethanol withdrawal. In addition, GIRK channels modulate long-term potentiation, a cellular mechanism of learning and memory. The goal of the current proposal is to assess the role of GIRK3 in fear-conditioned learning and memory and determine if changes in GIRK3 expression alter the withdrawal-induced changes in fear conditioning responses. The long-term goals of this project also include investigating the role of GIRK3 in individual learning related brain regions. In Aim 1, GIRK3 KO, HET, and wildtype (WT) littermates will be trained using two fear conditioning paradigms (delay fear conditioning and trace fear conditioning) in acute ethanol withdrawn and control animals. These two forms of conditioned learning are thought to utilize distinct but overlapping neural substrates, allowing us to assess the effect of reduced GIRK3 expression in different learning-related brain regions. After training, animals will be assessed for freezing in response to the training context and the conditioned stimulus. In Aim 2, animals will be trained following chronic ethanol withdrawal using these two fear conditioning types, allowing us to compare the effects of GIRK3 expression on learning and memory across multiple forms of ethanol withdrawal. These aims will not only determine the role of GIRK3-containing channels in fear conditioned learning and memory, but also characterize the effect of alcohol withdrawal on learning and memory and the extent of overlap between the learning effects and the withdrawal-reducing effects of GIRK3. The third aim will use the results from Aims 1 and 2 to identify specific brain regions that contribute to the altered withdrawal severity and the learning/memory changes observed in these GIRK3 genotypes. We will use RNAi to knock-down GIRK3 expression in individual brain regions of WT mice and assess the ability of the knock-down to attenuate ethanol withdrawal and related changes in learning and memory. Overall, this work will contribute to our understanding of how ethanol alters learning and memory systems, a key aspect of addiction development and relapse. PUBLIC HEALTH RELEVANCE: The development of alcohol addiction and the high rate of relapse in recovering alcoholics have been linked to alcohol-induced changes in learning and memory systems. While both learning/memory and alcohol addiction are complex processes that result from multiple genetic and environmental factors, several groups have reported overlapping contributing chromosomal regions for these two groups of behaviors. This project uses a novel genetic model of alcohol withdrawal to investigate withdrawal-induced changes in fear-based learning and memory.